Hypoxia is important in cardiovascular disease and cancer, the two leading causes of death in the US. Studies of these and other diseases need a tool for following changes in tissue hypoxia. We propose the HypoxyprobeTM system of markers can meet this need. HypoxyprobeTM markers have advantages of high water solubility, chemical stability, short plasma half-life, universal tissue distribution, efficient tissue uptake, low human toxicity in single and repeated injections and high marker adduct stability in tissue. Monoclonal antibodies will be prepared against seven HypoxyprobeTM analogues using techniques already established in our laboratories. Antibody reagents will be screened in ELISA assays. Sensitivity of HypoxyprobeTM analogues to hypoxia will be determined in tissue and multicellular spheroid cultures. Promising analogues will be tested in rodent and canine tumors in a STTR Phase II study. Leading experts believe that fluctuating hypoxia is an important factor in the pathophysiology of both normal and tumor tissues. Interventions designed to offset the damaging effects of fluctuating hypoxia are under investigation and the present proposal will provide powerful tools to support studies in these important areas of medicine. PROPOSED COMMERCIAL APPLICATION: NPI, Inc has already commercialized Hypoxyprobe (TM)-1, the lead compound in the Hypoxyprobe (TM) series of markers. It is expected that Hypoxyprobe(TM) analogues and associated monoclonal antibodies will also be widely used in the study of normal and tumor tissue hypoxia.